Busulfan [1,4-bis-(methanesulfonoxyl)butane], is a bifunctional alkylating agent which was first described by Haddow and Timmis (1953). Since the demonstration of its potent antitumor effects, it has been used extensively for treatment of malignant disease, especially hematologic malignancies and myeloproliferative syndromes (Galton, 1953; Ambs et al., 1971; Abe, 1975; Canellos, 1985; Hughes and Goldman, 1991; Collis, 1980). Its use was for long time limited to low dose oral therapy with palliative intent and frequent monitoring of the blood counts was routinely recommended (Canellos, 1985; Hughes and Goldman, 1991; Collis, 1980). The advent of some 2 to 3% of the patients developing busulfan-induced pulmonary fibrosis (Collis, 1980; Koch and Lesch, 1976; Oakhill et al., 1981), as well as occasionally severe, sometimes even irreversible myelosuppression after prolonged administration effectively deterred dose escalation beyond 8-10 mg daily (Canellos, 1985; Hughes and Goldman, 1991; Ganda and Mangalik, 1973; Albrecht et al., 1971).
In 1974, however, Santos and Tutschka investigated the use of busulfan to create a murine model of aplastic leukemia (Santos and Tutschka, 1974; Tutschka and Santos, 1975). Subsequently, the experience gained in this model system was used to introduce high-dose combination chemotherapy based on oral busulfan for pretransplant-conditioning of primates (Buckner, 1975), and subsequently patients undergoing both autologous and allogeneic marrow transplantation (Santos et al., 1983; Lu et al., 1984; Yeager et al., 1986; Tutschka et al., 1987; Peter et al., 1987; Copelan et al., 1989; Geller et al., 1989; Grochow et al., 1989; Sheridan et al., 1989). Since then, high dose busulfan, most commonly in combination with cyclophosphamide, has proven to be a most effective antileukemic regimen when used in conjunction with autologous or allogeneic hematopoietic stem cell support. A recent comparison between busulfan/cyclophosphamide (BuCy) and cyclophosphamide (Cy) combined with total body irradiation (TBI) for preparation of patients with hematologic malignancies undergoing allogeneic marrow transplantation illustrated that the BuCy regimen was well tolerated and at least as effective as the TBI-based regimen (Miller et al., 1991; Buckner et al., 1992; Schwertfeger et al.; 1992).
High-dose busulfan therapy has several advantages for use in marrow ablation/pretransplant treatment. First, when using chemotherapy alone for conditioning of patients undergoing marrow transplantation, one avoids the dependence on a radiation unit with, usually, limited capacity to deliver the necessary treatment on a fixed schedule. Second, high total radiation doses are very toxic, especially to the lungs, and may require special protective measures (shielding). Such excessive toxicity is usually not seen with combination chemotherapy. Third, a radiation based regimen can only be delivered to patients who have not been previously irradiated. Many patients with lymphoma, Hodgkin's disease and leukemia have had previous (extensive) radiation for control of locally aggressive disease in sanctuary sites like the central nervous system or to sites of bulky disease such as the mediastinum or the neck. Additional radiation as part of the pretransplantation conditioning regimen may cause irreversible and often fatal toxicity in such cases. However, a majority of previously radiated patients can safely receive a busulfan-based regimen, provided that the previous acute radiation toxicity (usually within the first 2-4 months after therapy) has subsided. Fourth, in selected patients who suffer recurrent leukemia after allogeneic marrow grafting, a second marrow transplant may still offer a chance for long-term disease control or even cure (Vaughn et al., 1991; Champlin et al., 1985; Sanders et al., 1988; Blume et al., 1987). Due to subclinical (irreversible) toxicity, a TBI-based regimen can only be utilized once in a patient's life time, whereas combination chemotherapy can be employed following a previous TBI-regimen. Busulfan-based chemotherapy will, therefore, serve as a valid alternative.
Oral busulfan has, unfortunately, several serious shortcomings. Thus, when used in high dose combinations with cyclophosphamide (and possibly additional chemotherapeutic agents), serious side effects in the liver and lungs are often encountered (Collis, 1980; Koch et al., 1976; Santos et al., 1983). Thus, several investigators have reported veno-occlusive disease (VOD) of the liver, leading to fatal liver failure, as the most serious side effect (Yeager et al., 1986; Geller et al. 1989; Grochow et al., 1989; Miller et al., 1991). Neurological disturbances like grand mal seizures and severe nausea and vomiting are also frequently encountered (Grigg et al., 1989; Marcus et al., 1984; Martell et al., 1987; Sureda et al., 1989; Vassal et al., 1990). It is impossible to predict which patients will develop liver failure, and it is further unknown whether the liver failure is due to toxicity from the systemic busulfan or whether it is mainly due to a first-pass phenomenon when busulfan is absorbed from the intestinal tract. Based on the somewhat sketchy information that is available on busulfan pharmacokinetics, it appears however, that patients who absorb a large fraction of the ingested dose, with a prolonged high busulfan plasma concentration, will be at increased risk for developing serious side effects (Marcus et al. 1984; Vassal et al., 1990). Another disadvantage with oral busulfan is, that patients who develop severe nausea and vomiting shortly (within 1-2 hours) after a dose has been delivered, will lose part of or the entire dose, and it may be virtually impossible to accurately determine how much of the dose has been lost in a vomiting subject. Further, the intestinal resorption of any delivered drug may be influenced by the patient's nutritional state, and by concurrent administration of other drugs affecting the intestinal microenvironment, as well as by whether the patient has eaten in close proximity to ingestion of the administered drug dose and, finally, by the inherent biological variability in intestinal absorption between different patients (Benet et al., 1985). Due to these uncertainties, oral administration of high-dose busulfan carries with it an inherent safety problem both from the potential danger of inadvertent overdosing with a risk for (lethal) toxicities, as well as from the hazard of (suboptimal) underdosing the patient with an inadvertently high potential for recurrent or persistent malignancy after the marrow transplant.
The in vivo distribution of busulfan labeled with the positron-emitting radionuclide carbon 11 was investigated in cynomolgus monkeys and in a human patient using positron emission tomography (Hassan et al., 1992). Radiotracer amounts of 11C-busulfan in a saline solution containing 10% ethanol were injected as an i.v. bolus. The concentration of busulfan was not reported but was likely insignificant compared to therapeutic levels. M Hassan has indicated to the inventor that the total dose injected was estimated at 1-2 .mu.g.
Giles et al. (1984) reportedly used busulfan to induce platelet dysfunction in rabbits by an intraperitoneal injection of busulfan dissolved in polyethyleneglycol at a dose of 60 mg/kg. The concentration of the solution is not given, and due to the slow solubilization of the busulfan, the authors heated the mixture excessively to promote mixing. This may have caused significant chemical degradation of the busulfan. Furthermore, busulfan given intraperitoneally is very toxic and causes significant local tissue damage.
An abstract of Kitamura's article (Kitamura, et al., 1979) relates to the well-known use of busulfan and cyclophosphamide which are typically given orally. The .sup.32 P is reported to have been administered intravenously.
In a study of bone marrow transplantation in the busulfan-treated rat, Tutschka and Santos reportedly injected busulfan prepared in 2.5% carboxymethylcellulose in water i.p. This injection also causes significant local tissue damage.
To circumvent the above shortcomings and hazards from oral administration of busulfan for chemotherapy with myeloablative intent, a chemically stable busulfan formulation that can be safely administered parenterally, i.e., via the intravenous (i.v.) route is needed.